In vitro assessment of the capacity of certain mycotoxin binders to adsorb some amino acids and water-soluble vitamins.

The objective of this study was to evaluate the capacity of 6 mycotoxin binders (MTB) to adsorb 3 AA and 4 water-soluble vitamins (WSV). Two experiments were conducted in in vitro conditions to simulate postruminal digestion with pepsin, malic acid, citric acid, acetic acid, and lactic acid at pH 3.0 and intestinal digestion with bile salts and pancreatin extract at pH 6.5. Experiment 1 was conducted with AA, and experiment 2 was conducted with WSV. Within experiment, main factors were the MTB (bentonite, clinoptiolite, sepiolite, montmorillonite, activated carbon, and yeast cell walls), the substrate (AA: Lys, Met, and Thr; WSV: B1, B2, B3, and B6), and the incubation strategy (substrates alone or mixed). Data were analyzed for the effects of main factors and their interactions. In experiment 1, the adsorption average for AA when incubated separately was 44.3%, ranging from 62.4% for Thr by clinoptiolite to 20.0% for Thr by activated carbon. When incubated together, the average adsorption was reduced to 19.9%, suggesting competition among substrates for adsorption. Adsorption ranged from 29.8% for Thr by yeast cell walls to 5.6% for Met by clinoptiolite, but there were significant interactions among MTB and AA. In experiment 2, the average adsorption of WSV when incubated separately or together was 34.1 and 45.1%, respectively, suggesting possible synergies among substrates. When vitamins were incubated separately, adsorption ranged from 90.5% for vitamin B1 to 4.0% for vitamin B3 by montmorillonite. Vitamins B1 (except by yeast cell walls) and B6 (except by bentonite, sepiolite, and montmorillonite) were absorbed the most, and vitamin B3 was absorbed the least (except by activated carbon and yeast cell walls, which were least together with vitamin B2). When vitamins were incubated together, adsorption ranged from 97.0% for vitamin B1 by montmorillonite to 0% for vitamin B2 by activated carbon and vitamin B3 by bentonite. Vitamins B1 by all MTB and B6 by clinoptiolite, sepiolite, and yeast cell walls were the most adsorbed, and vitamin B3 (except by activated carbon and yeast cell wall) was the least absorbed. There were significant interactions among MTB and WSV. Mycotoxin binders have a high degree of adsorption of the AA and WSV tested in in vitro conditions, which may limit their bioavailability. Results also suggest that when substrates were incubated together some interactions for adsorption occurred, which were competitive among AA and synergic among vitamins.

Effect of Diet Supplementation with the Mycotoxin Binder Montmorillonite on the Bioavailability of Vitamins in Dairy Cows.

The objective of this study was to determine the effect of the mycotoxin binder montmorillonite (MMT) supplemented in the diet of dairy cows on the bioavailability of vitamins A, D, E, B1 and B6. Six multiparous Holstein-Friesian cows were used in a crossover design with two periods. Treatments were a control diet with or without MMT. Vitamins were infused individually into the abomasum through the ruminal cannula. Blood samples were collected from the jugular vein at 0, 1, 2, 3, 4, 6, 9, 12, 24 and 48 h after the administration of each vitamin. Results showed that vitamin A reached maximal concentration (Tmax) at 5.3 h after dosing, the maximal concentration (Cmax) was 1.2 times higher than the basal concentration (Cbasal), and the area under the curve (AUC) was 739 arbitrary units. Vitamin B6 reached the Tmax at 13 h after dosing, the Cmax was 1.4 times higher than the Cbasal, and the AUC was 222 arbitrary units. No differences were observed in Cbasal, Tmax, Cmax and AUC of vitamin A and B6 between control vs. MMT-supplemented cows. Plasma concentrations of vitamins D, E and B1 had no concentration peaks, and were not affected by MMT addition. The lack of a response suggests that their plasma concentration may be tightly regulated. Results of this study do not show evidence that MMT affects the bioavailability of vitamins A and B6 in vivo.

Testing the protective effects of cyclodextrins vs. alternariol-induced acute toxicity in HeLa cells and in zebrafish embryos.

Alternariol (AOH) is a mycotoxin produced by Alternaria fungi, it appears as a contaminant in tomatoes, grains, and grapes. The chronic exposure to AOH may cause carcinogenic and xenoestrogenic effects. Cyclodextrins (CDs) are cyclic oligosaccharides, they form host-guest complexes with apolar molecules. In this study, the interactions of AOH with CD monomers and polymers were examined employing fluorescence spectroscopy. Thereafter, the protective effects of certain CDs vs. AOH-induced toxicity were investigated on HeLa cells and on zebrafish embryos. Our major observations are the following: (1) Sugammadex forms highly stable complex with AOH (K = 4.8 ×104 L/mol). (2) Sugammadex abolished the AOH-induced toxicity in HeLa cells, while native ß-CD did not show relevant protective effect. (3) Each CD tested decreased the AOH-induced mortality and sublethal adverse effects in zebrafish embryos: Interestingly, native ß-CD showed the strongest protective impact in this model. (4) CD technology may be suitable to relieve AOH-induced toxicity.

The efficacy of mycotoxin binders to control mycotoxins in feeds and the potential risk of interactions with nutrient: a review.

Mycotoxicosis are a common problem in livestock, where a group of six major mycotoxins represents a high risk for animal health and production profits. Mycotoxin binders (MTB) can reduce the mycotoxin burden in the gastrointestinal tract of the animal. Mycotoxin binders are classified in inorganic, as clays and activated carbon (AC), and organic, as yeast cell wall (YCW) and micro-ionized fibers. The adsorption of mycotoxins into MTB is due to: 1) chemical interactions where the cation exchange capacity involves different types of bounds like ion-dipole, Van der Walls forces, or hydrogen bonds; and 2) to physical characteristics of MTB like pore size, or mycotoxin structure and shape. The adsorption capacity of MTB is determined using different in vitro tests that mimic the gastrointestinal tract of the animals. A literature search was conducted to identify in vitro research where the efficacy of adsorption of MTB was determined. The search was based on 8 MTB [AC, bentonite, clinoptilolite, hydrated sodium calcium aluminosilicate (HSCAS), montmorillonite (MMT), sepiolite, YCW and zeolite] and 6 mycotoxins [aflatoxin (AF), deoxynivalenol (DON), fumonisin (FUM), ochratoxin (OTA), T-2 toxin and zearalenone (ZEA)]. Sixty-eight papers with 1842 data were selected and analyzed with the PROC MIXED of SAS. The response variable was the percentage mycotoxins adsorption by MTB, and the model included the fixed effects of MTB, mycotoxins, incubation media, pH and their interactions, and the random effect of the study. Differences were considered significant when P < 0.05 and with tendency when 0.05 < P < 0.10. The mycotoxins adsorption capacity was 83% ± 1.0 for AC, 76% ± 3.1 for MMT, 62% ± 1.0 for bentonite, 55% ± 1.9 for HSCAS, 52% ± 9.1 for sepiolite, 52% ± 4.3 for clinoptilolite and 44% ± 0.4 for YCW. For mycotoxins, the adsorption of AF was 76% ± 0.6, for FUM was 50% ± 1.8, for OTA was 42% ± 1.0, for ZEA was 48% ± 1.1, for DON was 35% ± 1.6, and for T-2 was 27% ± 2.8. The pH affected the adsorption capacity of YCW with higher adsorption at low pH, and the adsorption of OTA and ZEA, where OTA adsorption tended to be lower at intermediate pH, and adsorption of ZEA tended to be higher at the two-steps pH. The potential adsorption of some essential nutrients, including amino acids and vitamins, should also be considered. Results should be used as a guide in the selection of the appropriate mycotoxin binder based on the predominant mycotoxin in feeds.

Animal feeds are highly susceptible to mycotoxin contamination during harvest, storage, and processing. Mycotoxin binders represent an effective strategy to prevent mycotoxicosis in animals when supplemented into the diet. The efficacy of adsorption depends on the type of binder (inorganic or organic) and the physico-chemical properties of binders and mycotoxins. Data reviewed from the literature indicates that activated carbon has the highest adsorption capacity among different binders, and aflatoxins is the most adsorbed mycotoxin. However, the unspecific nature of the binding process results in some essential nutrients being also adsorbed.

Can Red Yeast (Sporidiobolus pararoseus) Be Used as a Novel Feed Additive for Mycotoxin Binders in Broiler Chickens?

Mycotoxin-contaminated feeds may negatively affect broiler chickens' health; hence, a sustainable approach to achieve mycotoxin elimination is necessary. This study aimed to evaluate the efficacy of red yeast (Sporidiobolus pararoseus; RY) as a novel mycotoxin binder in broilers. A total of 1440 one-week-old male broiler chicks were randomly assigned to 12 treatments in a 3 × 4 factorial design. The dietary treatments included three levels of mycotoxin-contaminated diets (0 µg kg−1 (0% of mycotoxin; MT), 50 µg kg−1 (50% MT), and 100 µg kg−1 (100% MT)) and four levels of mycotoxin binders (0.0 and 0.5 g kg−1 commercial binder, and 0.5 and 1.0 g kg−1 RY). Experimental diets were contaminated with aflatoxin B1, zearalenone, ochratoxin A, T-2 toxin, and deoxynivalenol in the basal diet. Furthermore, the parameters including feed intake, body weight, and mortality rate were recorded on a weekly basis. After feeding for 28 days, blood and organ samples were collected randomly to determine the blood biochemistry, relative organ weights, and gut health. The results indicated that mycotoxin-contaminated diets reduced the average daily weight gain (ADG), villus height (VH), and villus height per the crypt depth ratio (VH:CD) of the intestine, as well as the population of Lactobacillus sp. and Bifidobacterium sp. in the cecal (p < 0.05), whereas they increased the mycotoxins concentration in the blood samples and the apoptosis cells (TUNEL positive) in the liver tissue (p < 0.01) of broiler chicken. In contrast, RY-supplemented diets had better ADG values and lower chicken mortality rates (p < 0.05). Moreover, these combinations positively impacted the relative organ weights, blood parameters, bacteria population, intestinal morphology, and pathological changes in the hepatocytes (p < 0.05). In conclusion, RY supplementation effectively alleviated the toxicity that is induced by AFB1 and OTA, mainly, and could potentially be applied as a novel feed additive in the broiler industry.

Testing the extraction of 12 mycotoxins from aqueous solutions by insoluble beta-cyclodextrin bead polymer.

Mycotoxins are toxic metabolites of filamentous fungi; they are common contaminants in numerous foods and beverages. Cyclodextrins are ring-shaped oligosaccharides, which can form host-guest type complexes with certain mycotoxins. Insoluble beta-cyclodextrin bead polymer (BBP) extracted successfully some mycotoxins (e.g., alternariol and zearalenone) from aqueous solutions, including beverages. Therefore, in this study, we aimed to examine the ability of BBP to remove other 12 mycotoxins (including aflatoxin B1, aflatoxin M1, citrinin, dihydrocitrinone, cyclopiazonic acid, deoxynivalenol, ochratoxin A, patulin, sterigmatocystin, zearalanone, α-zearalanol, and ß-zearalanol) from different buffers (pH 3.0, 5.0, and 7.0). Our results showed that BBP can effectively extract citrinin, dihydrocitrinone, sterigmatocystin, zearalanone, α-zearalanol, and ß-zearalanol at each pH tested. However, for the removal of ochratoxin A, BBP was far the most effective at pH 3.0. Based on these observations, BBP may be a suitable mycotoxin binder to extract certain mycotoxins from aqueous solutions for decontamination and/or for analytical purposes.

The Efficacy of a Smectite-Based Mycotoxin Binder in Reducing Aflatoxin B1 Toxicity on Performance, Health and Histopathology of Broiler Chickens.

The aim of the experiment was to investigate the efficacy of a smectite-based clay binder (Toxo-MX) in reducing the toxicological effects of aflatoxin B1 (AFB1) in commercial broiler chickens. A total of 450 one-day old male broiler chickens were randomly allocated into three treatment groups with ten replicates of 15 birds each in a 42-day feeding experiment. The dietary treatments included a negative control (NC, a basal diet with no AFB1 and binder), a positive control (PC, a basal diet contaminated with 500 ppb of AFB1) and a smectite-based mycotoxin binder(Toxo-MX, PC with smectite clay binder). AFB1 challenge resulted in 14 to 24% depression in growth performance, elevated levels of aspartate aminotransferase (AST), and gamma-glutamyl transferase (GGT), organ enlargement and immuno-suppression.As compared to PC, feeding of Toxo-MX improved the final weight (15%; p < 0.0001), average daily gain (ADG) (15%; p < 0.001) and feed efficiency of broilers (13%; p < 0.0003) but did not have any effects on liver enzyme activities. Supplementation of smectite claysignificantly increased serum globulin levels and reduced the weight of the liver (p < 0.05) as compared to AFB1-fed broiler chickens. The severity of lesions (inflammatory and degenerative changes) observed in the liver, kidney, heart, pancreas, and lymphoid organs in PC birds was reduced by feeding smectite clay. The immuno-suppression caused by AFB1 was moderately ameliorated in Toxo-MX groupby stimulating the production of antibodies against IBD at day 42 (p < 0.05). In conclusion, dietary supplementation of a smectite-based mycotoxin binder to the diet containing AFB1 improved growth performance, reduced toxicological effects in liver and improved humoral immune response in broilers, suggesting its protective effect against aflatoxicosis.

Short Communication: Quantification of the Effect of Mycotoxin Binders on the Bioavailability of Fat-Soluble Vitamins In Vitro.

The aim of this study was to determine the capacity of six mycotoxin binders (MTBs) to adsorb vitamins A, D and E in an in vitro system that simulates gastric and intestinal digestion. Experiment 1 evaluated the recovery rate of vitamins A, D and E in the incubation conditions. In Experiment 2, the main factors were the MTB (bentonite, clinoptilolite, sepiolite, montmorillonite, active carbon and yeast cell walls), vitamins (A, D and E) and incubation type (vitamins incubated separately or together). The recovery was high for vitamin D (83%) and E (93%), but low for vitamin A (23%), for which no further analyses were conducted. When incubated separately, vitamin D was only adsorbed by yeast cell wall (20.2%). Vitamin E adsorption was highest with bentonite (54.5%) and montmorillonite (46.3%) and lowest with sepiolite (16.6%) and active carbon (18.5%). When incubated together, vitamin D was not adsorbed by any MTB. Vitamin E adsorption was highest in bentonite (61.8%) and montmorillonite (50.7%) and lowest in sepiolite (15.4%). Results indicate that the bioavailability of vitamin E, but not that of vitamin D, may be reduced in the presence of MTBs.

Impact of deoxynivalenol on intestinal explants of broiler chickens: An ex vivo model to assess antimycotoxins additives.

The impact of deoxynivalenol (DON) upon intestinal tissue of broilers was assessed by using jejunal explants in Ussing chambers and analyzing histopathological and immunohistochemical parameters; this system was also applied to evaluate the efficacy of an antimycotoxins additive (AMA). The explants were subjected to the following treatments within each experiment for 120 min: Experiment 1) T1 (control) - buffer solution, and T2 - 10 mg/L DON; and Experiment 2) T1 (control) - buffer solution, T2 - 10 mg/L DON, T3 - AMA (0.5%), and T4 - 10 mg/L DON + 0.5% AMA. In Experiment 1, DON triggered a reduction in the size of enterocytes as well as of their nuclei, an increase in cytoplasmic vacuolization and apical denudation of villi. Apoptotic cells count was also greater in DON-exposed explants. In Experiment 2, the AMA mitigated DON harmful effects; cytoplasmic vacuolization of enterocytes was reduced and the size of their nuclei was preserved. The additive also promoted a partial decrease in microvillus integrity, in size of enterocytes and in apoptotic cells count. The tested ex vivo model demonstrated the impact of DON upon the intestine as well as the efficacy of the AMA against its damaging effects.

Effect of a Mycotoxin Binder (MMDA) on the Growth Performance, Blood and Carcass Characteristics of Broilers Fed Ochratoxin A and T-2 Mycotoxin Contaminated Diets.

The present study was conducted to evaluate the efficacy of the feed additive, a novel multicomponent mycotoxin detoxifying agent (MMDA) containing modified zeolite (clinoptilolite), Bacillus subtilis, B. licheniformis, Saccharomyces cerevisiae cell walls, and silymarin, as detoxifiers of 0.5 mg/kg (0.5 ppm) ochratoxin A (OTA) and 1 mg/kg (1 ppm) T-2 toxin on broiler chickens. A total of 240 1-old broiler chickens (Ross 308) were randomly distributed into five different dietary treatments: (1) control (non-contaminated diet); (2) non contaminated diet + 3 g/kg of MMDA; (3) non-contaminated diet + 0.5 mg/kg OTA + 1 mg/kg T-2 toxin; (4) non-contaminated diet + 0.5 mg/kg OTA + 1 mg/kg T-2 toxin + 1 g/kg MMDA; and (5) non-contaminated diet + 0.5 mg/kg OTA + 1 g/kg T-2 toxin + 3 g/kg MMDA. The results showed that, in the starter period, from 1 to 10 days, the presence of OTA and T-2 mycotoxins reduced the consumption of feed and the growth of the broilers, and no effects of the detoxifying product were observed in the productivity of the chickens, at any of the doses tested, compared to the contaminated control (treatment 3). However, in the growing period, the same negative effect of mycotoxins was registered, but a recovery was observed in the consumption of feed and in the weight of the broilers that consumed 3 g/kg of the MMDA mycotoxin binder, reaching similar values to those of chickens fed uncontaminated control diets. The presence of mycotoxins in feed led to a reduction in the concentration of total proteins and albumin in blood compared to controls, and the presence of the detoxifying product partially reversed this effect. The breast yield of the chickens fed with mycotoxins was lower than that of the animals fed with the control feed and was not affected by the presence of the product tested, at 1 or 3 g/kg. The weight of the different organs (liver, gizzard, kidneys, or spleen), the intestinal pH, the histology of the small intestine, and oral lesions were not affected by the experimental treatments. In summary, the productive parameters and some blood and carcass characteristics of broiler chickens were impaired by the dietary presence of OTA and T-2 toxin. The tested product included at 1 or 3 g/kg feed in contaminated diets improved performance and seems to be effective in partly counteracting the deleterious effects of the tested mycotoxins.

Effectiveness of Training and Use of Novasil Binder in Mitigating Aflatoxins in Cow Milk Produced in Smallholder Farms in Urban and Periurban Areas of Kenya.

Aflatoxins, which commonly contaminate animal feeds and human food, present a major public health challenge in sub-Saharan Africa. After ingestion by cows, aflatoxin B1 is metabolized to aflatoxin M1 (AFM1), some of which is excreted in milk. This study involved smallholder dairy farms in urban and periurban areas of Nairobi and Kisumu, Kenya. The objective was to determine the effectiveness of training and providing farmers with aflatoxin binder (NovaSil®) on AFM1 contamination in raw milk. A baseline survey was undertaken and 30 farmers whose milk had AFM1 levels above 20 ppt were randomly selected for inclusion in the study. Of these, 20 farmers were part of the intervention, and were given training on the usage of the NovaSil® binder, while 10 served as a control group. All farmers were visited biweekly for three months for interviews and milk samples were collected to measure the AFM1 levels. The AFM1 levels were quantified by enzyme linked immunosorbent assay. The NovaSil® binder significantly reduced AFM1 concentrations in the raw milk produced by the farmers in the intervention group over the duration of the study (p < 0.01). The control farms were more likely to have milk with AFM1 levels exceeding the regulatory limit of 50 ppt compared to the intervention farms (p < 0.001) (odds ratio = 6.5). The farmers in the intervention group perceived that there was an improvement in milk yield, and in cow health and appetite. These farmers also felt that the milk they sold, as well as the one they used at home, was safer. In conclusion, the use of binders by dairy farmers can be effective in reducing AFM1 in milk. Further research is needed to understand their effectiveness, especially when used in smallholder settings.

Assessment of H-ß zeolite as an ochratoxin binder for poultry.

Most of the cereal-based ingredients used in poultry feed are contaminated with ochratoxin-A (OTA). We have investigated H-ß zeolite (HBZ) as a new OTA binder for poultry, along with widely used clay mineral-based product (CM), using in vitro and in vivo methods. In vitro binding experiment was carried out using a biphasic assay, consisting of adsorption at pH 3.2 and desorption at pH 6.8. High adsorption (>98%) with less desorption (<5%) was observed for HBZ, whereas CM showed high binding (>98%) and moderate desorption (48%). In the in vitro experiments with the different simulated gastro-intestinal pH buffers, HBZ did not desorb OTA at any of the pH. Desorption of OTA was observed with CM, as the pH increases. From the in vitro kinetic and chemisorption studies, faster, stronger, and higher adsorption was observed for HBZ. Thermodynamic studies showed positive entropy (22.76 KJ/mol K) for HBZ, signifying predominant hydrophobic interactions towards OTA, whereas CM exhibited negative entropy (-3.67 KJ/mol K). The in vivo binding efficacy of HBZ and CM was tested in 5-wk-old broiler chickens. The study consisted of 4 experimental groups, each with 6 replicates having 2 birds per replicate. The groups were control, negative control (no toxin binder), T1 (HBZ at 1 kg/ton of feed), and T2(CM at 1 kg/ton of feed). Except control, all the replicates received 20 µg of OTA in the feed. Excreta samples of T1, T2, and NC contained 11.57, 7.16, and 2.78 µg of OTA respectively, which was significantly different from each other (P < 0.05). A growth performance trial was conducted in broiler chickens for 35 D. A total of 288 one-day-old birds were randomly segregated to 3 treatment groups, each with 8 replicates of 12 birds each. Treatment groups consisted of control, T1, and T2, treated with no toxin binder, HBZ, and CM at 1 kg/ton of feed, respectively. None of the treatment groups including control, affected BW gain, and feed conversion ratio (P > 0.05).

Efficacy of a preparation of algae interspaced bentonite as a feed additive for all animal species.

Following a request from the European Commission, the EFSA Panel on Additives and Products or Substances used in Animal Feed (FEEDAP Panel) was asked to deliver a scientific opinion on the efficacy of a preparation of algae interspaced bentonite when used as aflatoxin B1 (AfB1) binder for all animal species; the additive is composed of bentonite feed grade and algae belonging to Ulva spp. The European Commission request follows an opinion of the FEEDAP Panel published in 2016; in that opinion, the FEEDAP Panel could not conclude on the efficacy of the additive. The applicant has submitted additional information to allow the FEEDAP Panel to complete its assessment; these additional data, related to the efficacy of the additive, are the subject of this opinion. In the efficacy study provided, the use of the additive reduced the AfM1 in milk, although this effect seemed to be not consistent over time; this study presented some weaknesses and limitations (incomplete data analysis, reporting and lack of determination of the active substance of the additive in feed) which do not allow the FEEDAP Panel to draw any conclusion on the efficacy of the additive. Due to insufficient evidence, the FEEDAP Panel cannot conclude on the efficacy of the algae interspaced bentonite as a technological feed additive for all animal species.

In vitro studies on gastrointestinal monogastric and avian models to evaluate the binding efficacy of mycotoxin adsorbents by liquid chromatography-tandem mass spectrometry.

OBJECTIVE: The objective of this study is evaluating the efficacies of 11 mycotoxin adsorbent products, marketed in South East Asia. Three prominently occurring mycotoxins; aflatoxin B1 (AFB1), deoxynivalenol (DON), and zearalenone (ZEN) were simultaneously spiked into the samples. MATERIALS AND METHODS: Samples were simultaneously tested in vitro in phosphate buffer and simulated at different pH conditions in the gastrointestinal tracts of the porcine and avian model, analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). RESULTS: All mycotoxin adsorbent products had high efficacy at over 90% for AFB1 adsorption in both GI porcine and avian models. AFB1 could be adsorbed more in acidic condition than the basic condition. ZEN adsorption was determined to be more stable at pH 3 than pH 6.5 or 8.4, in which pH condition might influence on ZEN desorption rate. DON was poorly adsorbed by all tested agents. CONCLUSIONS: The finding showed that the adsorption rate varied depending on the type of adsorbent. Our results might provide useful information regarding the efficacy of mycotoxin adsorbents commercially marketed in the region.

Cyclodextrins Can Entrap Zearalenone-14-Glucoside: Interaction of the Masked Mycotoxin with Cyclodextrins and Cyclodextrin Bead Polymer.

Zearalenone (ZEN) is a Fusarium-derived xenoestrogenic mycotoxin. In plants, zearalenone-14-O-ß-d-glucoside (Z14G) is the major conjugated metabolite of ZEN, and is a masked mycotoxin. Masked mycotoxins are plant-modified derivatives, which are not routinely screened in food and feed samples. Cyclodextrins (CDs) are cyclic oligosaccharides built up from D-glucopyranose units. CDs can form stable host-guest type complexes with lipophilic molecules (e.g., with some mycotoxins). In this study, the interaction of Z14G with native and chemically modified ß- and γ-CDs was examined employing fluorescence spectroscopy and molecular modeling. Furthermore, the removal of Z14G from aqueous solution by insoluble ß-CD bead polymer (BBP) was also tested. Our results demonstrate that Z14G forms the most stable complexes with γ-CDs under acidic and neutral conditions (K ≈ 103 L/mol). Among the CDs tested, randomly methylated γ-CD induced the highest increase in the fluorescence of Z14G (7.1-fold) and formed the most stable complexes with the mycotoxin (K = 2 × 103 L/mol). Furthermore, BBP considerably reduced the Z14G content of aqueous solution. Based on these observations, CD technology seems a promising tool to improve the fluorescence analytical detection of Z14G and to discover new mycotoxin binders which can also remove masked mycotoxins (e.g., Z14G).

Interactions of Mycotoxin Alternariol with Cyclodextrins and its Removal from Aqueous Solution by Beta-Cyclodextrin Bead Polymer.

Alternariol is an Alternaria mycotoxin that appears in fruits, tomatoes, oilseeds, and corresponding products. Chronic exposure to it can induce carcinogenic and xenoestrogenic effects. Cyclodextrins (CDs) are ring-shaped molecules built up by glucose units, which form host-guest type complexes with some mycotoxins. Furthermore, insoluble CD polymers seem suitable for the extraction/removal of mycotoxins from aqueous solutions. In this study, the interactions of alternariol with ß- and γ-CDs were tested by employing fluorescence spectroscopic and modeling studies. Moreover, the removal of alternariol from aqueous solutions by insoluble ß-CD bead polymer (BBP) was examined. Our major observations/conclusions are the following: (1) CDs strongly increased the fluorescence of alternariol, the strongest enhancement was induced by the native γ-CD at pH 7.4. (2) Alternariol formed the most stable complexes with the native γ-CD (logK = 3.2) and the quaternary ammonium derivatives (logK = 3.4-3.6) at acidic/physiological pH and at pH 10.0, respectively. (3) BBP effectively removed alternariol from aqueous solution. (4) The alternariol-binding ability of ß-CD polymers was significantly higher than was expected based on their ß-CD content. (5) CD technology seems a promising tool to improve the fluorescence detection of alternariol and/or to develop new mycotoxin binders to decrease alternariol exposure.

Effects of Feeding a Mycotoxin Binder on Nutrient Digestibility, Alkaloid Recovery in Feces, and Performance of Lambs Fed Diets Contaminated with Cereal Ergot.

As contamination with cereal ergot has been increasing in western Canada, this study evaluated impacts of feeding a mycotoxin binder (Biomin® II; BB) on nutrient digestibility, alkaloid recovery in feces, and lamb growth performance. Forty-eight ram lambs (25.9 ± 1.4 kg) were randomly assigned to one of four barley-based diets: Control (C), no added alkaloids, Control + BB fed at 30 g/head per day (CBB); Ergot, 2564 ppb total R + S epimers (E); Ergot + BB, 2534 ppb R + S epimers (EBB). Lambs were fed ab libitum for up to 11 weeks until slaughter at >46 kg live weight. Both average daily gain (ADG) and gain/feed ratio were greater (p < 0.01) for lambs fed C and CBB diets as compared with those containing added ergot, although dry matter intake was not affected by dietary ergot or BB. Serum prolactin concentrations were two times higher in EBB- compared with E-fed lambs (p < 0.05), although both were lower than in C or CBB (p < 0.001) lambs. Rectal temperatures were greater in lambs receiving dietary ergot (p ≤ 0.001) than in C- and CBB-fed lambs. In a digestibility study using eight ram lambs, treatment with BB increased neutral detergent fiber (NDF) digestibility (p = 0.01). Nitrogen retention (g) was greater (p < 0.05) for lambs receiving C or CBB compared with ergot-contaminated diets. Feces of EBB lambs had 38.5% greater (p < 0.001) recovery of alkaloids compared with those fed E. Based on sparing of prolactin, BB may reduce impacts of ergot alkaloids by increasing their excretion in feces. Accordingly, concentrations of dietary alkaloids, which would not harm sheep, would be increased by feeding BB.

In vitro model to assess the adsorption of oral veterinary drugs to mycotoxin binders in a feed- and aflatoxin B1-containing buffered matrix.

Mycotoxin binders are feed additives which are mixed in the feed to adsorb mycotoxins and thereby reducing their toxic effects on animals. Interactions with orally administered veterinary medicinal products, such as antimicrobials or coccidiostats, have been reported previously. This paper describes an in vitro model to screen the interaction between mycotoxin binders and veterinary drugs with respect to the non-specific binding of drugs. It is designed as a static setup using a single concentration of drug and binder in a feed-containing or a feed-plus-mycotoxin-containing matrix, buffered at different pH values. The model was applied to two frequently used antimicrobials in veterinary medicine, doxycycline (DOX) and tylosin (TYL), one major mycotoxin, aflatoxin B1 (AFB1), and four mycotoxin binders. Proportions of feed, DOX or TYL, AFB1, and binder are equivalent to the in vivo situation for broiler chickens, while pH and volume of the buffer are representative of the gastrointestinal tract of chickens. A substantial binding of DOX (~ 88%) and TYL (~ 66%) to the feed-matrix was observed. For the mycotoxin binders, similar results were obtained for DOX and TYL; more specifically up to an inclusion rate of 20 g binder/kg feed, no significant binding was demonstrated, determined as the free concentration of DOX and TYL. A single exception was noticed for TYL and one specific bentonite-based mycotoxin binder, for which no significant interaction could be demonstrated up to 10 g binder/kg but there was an effect at 20 g/kg. In all cases, there was no competition between the tested drugs DOX or TYL and the mycotoxin AFB1 for binding to the bentonite-based mycotoxin binder.

Safety and efficacy of bentonite as a feed additive for all animal species.

The EFSA Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) received a request from the European Commission to assess the safety and efficacy of bentonite when used as a technological feed additive (substances for reduction of the contamination of feed by mycotoxins) for all animal species. The applicant, EUBA aisbl (European Bentonite Association) representing six companies, submitted to EFSA a technical dossier to support the application. The applicant proposes to use bentonite at the maximum level of 20,000 mg/kg complete feed. The additive apparently interferes with the analysis of aflatoxin B1 in feed. The safety of the additive was already evaluated by the Panel in an opinion delivered in 2012. Bentonites are safe for all animal species, the consumers and the environment when used at a maximum level of 20,000 mg/kg complete feed. The results of a new genotoxicity study reinforced the previous conclusion that smectites are non-genotoxic. Bentonites are not skin irritants but might be mildly irritant to the eye; based on a new study submitted, the additive is not a skin sensitiser. Owing to its silica content, the additive is a hazard by inhalation for the users. The in vitro study showed that the di- and tri-octahedral smectites tested can adsorb aflatoxin B1 at different concentrations and at pH 5; however, no adequate in vivo studies were available. Therefore, the Panel cannot draw conclusions on the additive's efficacy. The Panel further considers the safety and efficacy conclusions to apply equally to the di- and tri-octahedral smectites under assessment. The FEEDAP Panel posted some recommendations regarding the maximum content of other minerals in the additive and the incompatibilities of the additive with other medicinal substances. The Panel also drew a remark concerning the denomination of the additive and the current regulatory definition of Bentonite.

Safety and efficacy of microorganism DSM 11798 as a technological additive for all avian species.

Biomin® BBSH 797 is the trade name for a feed additive containing viable cells of an unnamed bacterium (DSM 11798). The EFSA Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) produced an opinion on the safety and efficacy of this additive when used with pigs concluding that the additive itself did not raise any safety concern. It was confirmed that the additive could reduce the trichothecene, deoxynivalenol (DON), producing the less toxic de-epoxy metabolite. The applicant is now seeking authorisation for the use of the additive in feed for all avian species and, since the current authorisation only covers DON, the amending of this authorisation to include all trichothecene mycotoxins. Chickens and turkeys for fattening and laying hens showed no adverse effects when the additive was added to diets at 1000 times the recommended dose. Consequently, the additive is considered safe for these species/categories when used at the recommended dose of 1.7 × 108 colony-forming units (CFU)/kg complete feed. This conclusion is extended to all avian species. The use of the additive in feed for all avian species is not expected to introduce concerns for consumers, users or the environment not previously considered. In vitro and in vivo studies showed that the inclusion of the additive at the recommended dose of 1.7 × 108 CFU/kg was effective in reducing contaminating DON in feed when given to avian species with a concomitant production of the less toxic de-epoxy metabolite. The additive was shown to reduce the 12,13-epoxide group of representative trichothecenes. It would be reasonable to assume a similar reaction with other mycotoxins of the same structural type independent of the animal species or category receiving contaminated feed. The additive is compatible with the coccidiostats monensin sodium, salinomycin sodium, narasin, narasin/nicarbazin, nicarbazin, robenidine hydrochloride and diclazuril.